Subcutaneous pharmacokinetics of the cardiac hormone vessel dilator.

Vessel dilator, a hormone synthesized in the heart, eliminates 71% of human small-cell lung cancers and 67% of human breast cancers growing in mice when given subcutaneously (s.c.) via osmotic pumps. The pharmacokinetics of s.c. administered vessel dilator have not been evaluated previously. In the present study, the pharmacokinetics of vessel dilator following s.c. bolus (ScB) or 3 h s.c. infusion (ScI) were compared with those following i.v. bolus (IvB) administration in male Fischer 344 rats. The half-life (t½ ) of vessel dilator after ScI, IvB and ScB was 54, 43 and 30 min, respectively. The tmax for vessel dilator after IvB, ScB and ScI administration was 1.5, 23 and 156 min, respectively, whereas the corresponding Cmax values were 3749, 887 and 471 ng/L (normalized against the dose used for ScB and IvB). The area under the curve (AUC0-∞ ) for vessel dilator was 1166, 880 and 1652 ng h/mL (normalized) following IvB, ScB and ScI administration, respectively. The volume of distribution for vessel dilator was 2.38, 0.92 and 1.08 L following IvB, ScB and SCI administration, respectively; corresponding clearance values were 1.69, 1.50 and 0.78 L/h, respectively. Plasma concentrations of vessel dilator after each of the three methods of administration mirrored their predicted concentration-time profiles. We conclude that vessel dilator administered via ScI has a significantly greater AUC and t½ and slowed clearance compared with IvB or ScB administration (P < 0.001), suggesting that s.c. infusion is the preferred method of administration, based on pharmacokinetics, to treat cancers.

Reduction of leptin levels by four cardiac hormones: Implications for hypertension in obesity.

Circulating levels of leptin are increased in obesity and have been proposed to contribute to the development of hypertension in obese individuals. Four cardiac hormones, specifically, vessel dilator, long-acting natriuretic peptide (LANP), kaliuretic peptide and atrial natriuretic peptide (ANP), have blood pressure-lowering properties and correlate with the presence of hypertension in obesity. The objective of this study was to determine whether one or more of these cardiac hormones was able to decrease the levels of leptin in the hypothalamus, an area of the brain that has been demonstrated to synthesize more than 40% of leptin in the circulation. The effects of these four cardiac hormones on leptin were examined using dose-response curves in the rat hypothalamus, which synthesizes leptin. Vessel dilator, LANP, kaliuretic peptide and ANP maximally decreased the levels of leptin in hypothalamic cells by 79, 76, 80 and 62%, respectively (P<0.0001 for each). The cardiac hormones decreased leptin levels over a concentration range of 100 pM to 10 µM, with the most significant reductions in leptin levels occurring when the concentrations of the hormones were at micromolar levels. The results of the study suggest that the four cardiac hormones lead to significant reductions in hypothalamic leptin levels, which may be an important mechanism for alleviating leptin-induced hypertension in obesity.

Cardiac hormones target nuclear oncogenes c-Fos and c-Jun in carcinoma cells.

BACKGROUND: c-Fos is a cellular proto-oncogene which dimerizes with c-Jun proto-oncogene to form AP-1 transcription factor, which upregulates transcription of genes involved in proliferation and cancer formation. Four cardiac hormones, that is, long-acting natriuretic peptide (LANP), vessel dilator, kaliuretic peptide (KP) and atrial natriuretic peptide (ANP) with anticancer effects in vivo are potent inhibitors of the Ras-MEK 1/2-ERK 1/2 kinase cascade and signal transducer and activator of transcription-3 (STAT-3) that activate c-Fos and c-Jun. These four cardiac hormones were investigated for their effects on proto-oncogenes c-Fos and c-Jun within the nucleus of cancer cells. MATERIALS AND METHODS: Four cardiac hormones were evaluated for their ability to decrease proto-oncogenes c-Fos and c-Jun, measured by ELISA in extracted nuclei of three human cancer cell lines. RESULTS: Vessel dilator, LANP, KP and ANP over a concentration range of 100 pM-10 µM, maximally decreased c-Fos by 61%, 60%, 61% and 59% in human hepatocellular cancer cells, by 82%, 74%, 78% and 74% in small-cell lung cancer cells, and by 82%, 73%, 78% and 74% in human renal adenocarcinoma cells. c-Jun was maximally reduced by vessel dilator, LANP, KP and ANP by 43%, 31%, 61% and 35% in hepatocellular cancer cells, by 65%, 49%, 59% and 40% in small-cell lung cancer cells, and by 47%, 43%, 57% and 49% in renal cancer cells. CONCLUSION: Four cardiac hormones are potent inhibitors of c-Fos and c-Jun proto-oncogenes within the nucleus of cancer cells.

Potent selective inhibition of STAT 3 versus STAT 1 by cardiac hormones.

Signal transducers and activators of transcription (STATs) are the final "switches" that activate gene expression patterns that lead to human malignancy. Extracellular signal-regulated kinases (ERK 1/2) activate STAT 3; four cardiovascular hormones inhibit ERK 1/2 kinases, leading to the hypothesis that they may also inhibit STATs. These four cardiac hormones, i.e., vessel dilator, long-acting natriuretic peptide (LANP), kaliuretic peptide, and atrial natriuretic peptide (ANP), eliminate human cancers growing in mice. These four cardiac hormones' effects on STATs 1 and 3 were examined in human small-cell lung cancer and human pancreatic adenocarcinoma cells. Vessel dilator, LANP, kaliuretic peptide, and ANP maximally decreased STAT 3 by 88, 54, 55, and 65 %, respectively, at their 1 µM concentrations in human small-cell lung cancer cells and STAT 3 by 66, 57, 70, and 77 % in human pancreatic adenocarcinoma cells, respectively. The cardiac hormones (except LANP) also significantly decreased STAT 3 measured by Western blots. These cardiac hormones did not decrease STAT 1 in either human small-cell lung cancer or pancreatic adenocarcinoma cells. We conclude that these four cardiac hormones are significant inhibitors of STAT 3, but not STAT 1, in human small-cell lung cancer and pancreatic adenocarcinoma cells, which suggests a specificity for these hormones' anticancer mechanism(s) of action enzymology in human cancer cells.

Novel dual inhibitors of vascular endothelial growth factor and VEGFR2 receptor.

BACKGROUND: Vascular endothelial growth factor (VEGF) helps control tumour growth via causing new capillaries growth in tumours. Four cardiac hormones [i.e. vessel dilator, long-acting natriuretic peptide (LANP), kaliuretic peptide (KP) and atrial natriuretic peptide (ANP)] that eliminate up to up to 86% of human small-cell lung cancers growing in mice were investigated for their effects on VEGF and the VEGFR2/KDR/Flk-1 receptor. The VEGFR2 receptor is the main receptor mediating VEGF's cancer-enhancing effects. MATERIALS AND METHODS: Four cardiac hormones were evaluated for their ability to decrease VEGF/VEGFR2 measured by ELISAs in three human cancer cell lines. RESULTS: Vessel dilator, LANP, KP and ANP, over a concentration range of 100 pM to 10 µM, maximally decreased the VEGFR2 receptor in human pancreatic adenocarcinoma cells by 48%, 49%, 74% and 83%. Vessel dilator, LANP, KP and ANP decreased the VEGFR2 receptor by 77%, 89%, 88% and 67% in human small-cell lung cancer cells and by 48%, 92%, 64% and 71% in human prostate cancer cells. These results were confirmed with the cardiac hormones also decreasing the VEGFR2 receptor measured by Western blots. VEGF itself in pancreatic carcinoma cells was decreased by 42%, 58%, 36% and 40% by vessel dilator, LANP, KP and ANP. VEGF levels were decreased 25%, 23%, 17% and 23% in small-cell lung cancer cells and decreased by 24%, 20%, 23% and 24% in prostate cancer cells by vessel dilator, LANP, KP and ANP. CONCLUSION: Four cardiac hormones are the first dual inhibitors of VEGF and the VEGFR2/KDR/Flk-1 receptor.

Cardiac hormones are c-Jun-N-terminal kinase 2-inhibiting peptides.

BACKGROUND: Four cardiac peptide hormones, namely vessel dilator, long-acting natriuretic peptide (LANP), kaliuretic peptide, and atrial natriuretic peptide (ANP) have anticancer effects. MATERIALS AND METHODS: The effects of these four cardiac hormones on human c-Jun-N-terminal kinase 2 (JNK2) were examined in human small cell lung cancer and human prostate cancer cells. RESULTS: Vessel dilator, LANP, kaliuretic peptide and ANP maximally reduced expression of JNK2 by 89%, 56%, 45%, and 28%, respectively (each at p<0.0001) in human small cell lung cancer cells. In human prostate adenocarcinoma cells, JNK2 was maximally decreased 76%, 56%, 45%, (each at p<0.0001), and 28% (p<0.01) secondary to vessel dilator, LANP, kaliuretic peptide and ANP, respectively. CONCLUSION: These results indicate that four cardiac hormones are significant inhibitors (by up to 89%) of JNK2 in human small cell lung cancer cells and up to 76% in human prostate adenocarcinoma cells as part of their anticancer mechanism(s) of action.

Dual pressure from antiretroviral therapy and cell-mediated immune response on the human immunodeficiency virus type 1 protease gene.

Human immunodeficiency virus (HIV)-specific CD8(+) T-lymphocyte pressure can lead to the development of viral escape mutants, with consequent loss of immune control. Antiretroviral drugs also exert selection pressures on HIV, leading to the emergence of drug resistance mutations and increased levels of viral replication. We have determined a minimal epitope of HIV protease, amino acids 76 to 84, towards which a CD8(+) T-lymphocyte response is directed. This epitope, which is HLA-A2 restricted, includes two amino acids that commonly mutate (V82A and I84V) in the face of protease inhibitor therapy. Among 29 HIV-infected patients who were treated with protease inhibitors and who had developed resistance to these drugs, we show that the wild-type PR82V(76-84) epitope is commonly recognized by cytotoxic T lymphocytes (CTL) in HLA-A2-positive patients and that the CTL directed to this epitope are of high avidity. In contrast, the mutant PR82A(76-84) epitope is generally not recognized by wild-type-specific CTL, or when recognized it is of low to moderate avidity, suggesting that the protease inhibitor-selected V82A mutation acts both as a CTL and protease inhibitor escape mutant. Paradoxically, the absence of a mutation at position 82 was associated with the presence of a high-avidity CD8(+) T-cell response to the wild-type virus sequence. Our results indicate that both HIV type 1-specific CD8(+) T cells and antiretroviral drugs provide complex pressures on the same amino acid sequence of the HIV protease gene and, thus, can influence viral sequence evolution.

Identification of epitope-like consensus motifs using mRNA display.

The mRNA display approach to in vitro protein selection is based upon the puromycin-mediated formation of a covalent bond between an mRNA and its gene product. This technique can be used to identify peptide sequences involved in macromolecular recognition, including those identical or homologous to natural ligand epitopes. To demonstrate this approach, we determined the peptide sequences recognized by the trypsin active site, and by the anti-c-Myc antibody, 9E10. Here we describe the use of two peptide libraries of different diversities, one a constrained library based on the trypsin inhibitor EETI-II, where only the six residues in the first loop were randomized (6.4 x 10(7) possible sequences, 6.0 x 10(11) sequences in the library), the other a linear-peptide library with 27 randomized amino acids (1.3 x 10(35) possible sequences, 2 x 10(13) sequences in the library). The constrained library was screened against the natural target of wild-type EETI, bovine trypsin, and the linear library was screened against the anti-c-myc antibody, 9E10. The analysis of selected sequences revealed minimal consensus sequences of PR(I,L,V)L for the first loop of EETI-II and LISE for the 9E10 epitope. The wild-type sequences, PRILMR for the first loop of EETI-II and QKLISE for the 9E10 epitope, were selected with the highest frequency, and in each case the complete wild-type epitope was selected from the library.

The 2003 anatomy ceremony: a service of gratitude.

In keeping with a long-standing tradition, Yale Medical and Physician Associate students gather at a ceremony each year after the completion of the anatomy course. The ceremony is a chance to reflect and to give thanks. It gives students the opportunity to articulate their gratitude to the selfless individuals who donated their bodies for the benefit of education. Many family members of the donors attend the ceremony. By reading poetry, performing musical pieces, and presenting works of art, the students and their teachers express some of the emotions and thoughts that the anatomy course has evoked. The following are some of the contributions presented at this year's ceremony.